P987: IRAK1 AND JAK2 DUAL INHIBITION WITH PACRITINIB PREVENTS MYELOFIBROSIS-LIKE PHENOTYPE IN AN INFLAMMATION-DRIVEN MURINE MODEL
Authors: Ernesto José Cuenca Zamora, Pedro Jesús Guijarro Carrillo, María José López Poveda, María Luz Morales Fernández, Maria L Lozano, Rocío González-Conejero, Constantino Martínez, Raúl Teruel Montoya, Francisca Ferrer-Marín
Journal: HemaSphere (2023)

Topic: 15. Myeloproliferative neoplasms - Biology & Translational Research Background: Inflammation plays a key role in the development and progression of myeloproliferative neoplasms (MPN). MPN patients show elevated plasma levels of proinflammatory cytokines, mainly due to activation of the NF-κB pathway. It has been described that miR-146a regulates the NF-κB pathway by inhibiting IRAK1/TRAF6. Indeed, miR-146a-/- mice develop a myelofibrosis (MF)-like phenotype with aging. We have previously shown that the germline variant rs2431697, associated with lower expression levels of MIR146A in humans, predicts early progression to secondary MF (Leukemia 2020). We have also shown in a miR-146a-/- (KO) mouse model without driver mutations that these mice exhibit an increase in STAT3 signaling with aging, which likely contributes to the MF phenotype. Pacritinib is a JAK1-sparing JAK2/IRAK1 inhibitor used for the treatment of MF. The impact of a dual IRAK1/JAK2 inhibitor on an inflammation driven MF phenotype has not been previously investigated. Aims: To evaluate whether NF-κB signaling inhibition may suppress chronic inflammation and prevent the development of myeloproliferation and fibrosis using an inflammation model lacking driver mutations: a mouse deficient for miR-146a. Methods: Three-month-old KO mice were ad libitum treated with either 3 (3m) or 6 months (6m) of chow with pacritinib at 250 mg/kg/day. In addition, 3m KO and wild-type (WT) mice fed under the same conditions but on pacritinib-free chow (“untreated”) were used as controls (n=8-10 mice/group). After 3 or 6 months, in 6 or 9-month-old mice respectively, we evaluated: cell counts in peripheral blood; in spleen: size, weight, architecture (white/red pulp ratio), extramedullary hematopoiesis (number and percent of dysplastic megakaryocytes), and expression of STAT3, pTyr-STAT3, STAT5, IRAK1 and p65 by western blot (WB); and in bone marrow (BM): histology. Results: Consistent with the miR-146a-/- phenotype, KO compared to WT mice, at both ages, showed cytopenias; large spleen with architecture loss and foci of extramedullary hematopoiesis (Fig. 1A); as well as reticulin fibrosis with osteosclerosis in BM (Fig. 1B). Due to the spleen architecture loss, we found undetectable levels of STAT5, IRAK1 and p65 but slightly higher of pTyr-STAT3 and STAT3 in KO (vs WT) mice. After 3 or 6 months of pacritinib treatment, KO mice (vs untreated KO) showed a significant increase in platelets (3m: p<0.01; 6m: p<0.0001) without worsening of anemia, lower spleen weight (3m: 1.4-fold, p<0.05; 6m: 7.1-fold, p<0.0001), better preservation of spleen architecture (3m, p<0.001; 6m, p<0.05) and fewer foci of extramedullary hematopoiesis (3m, p<0.01; 6m, p<0.001, Fig. 1A). Following the recovery of spleen architecture, a partial normalization of the levels of all proteins examined by WB was observed, with a tendency to resemble WT mice profile. Histological analysis of BM revealed that dual inhibition of JAK2/IRAK1 signaling with pacritinib for 3 or 6 months was able to prevent reticulin fibrosis (p<0.001) and osteosclerosis (p<0.05) (Fig. 1B), observed in untreated KO mice. Summary/Conclusion: In this in vivo MF-like model, particularly useful to study the inflammatory component of MF without driver mutations, dual JAK2+IRAK1 inhibition, with inhibitors such as pacritinib, prevents the development of thrombocytopenia, splenomegaly, extramedullary hematopoiesis and BM fibrosis, postulated as a potential modifier of the natural course of these diseases. The improvement of inflammatory symptoms in MF patients receiving pacritinib, which does not inhibit JAK1, could be explained by inhibition of the NF-κB pathway through IRAK1.Keywords: Bone Marrow Fibrosis, Myeloproliferative disorder, Myelofibrosis, NF- B